水稻LPA2基因调控分蘖角度的分子机理研究

分蘖角度作为水稻株型的重要组成部分，是决定产量的关键因素之一，与地上部分重力反应密切相关。前期在贵州平塘黑糯EMS突变体库中筛选到一株型松散、分蘖角度增大且地上部分重力反应降低的突变体lpa2。通过基因定位克隆了LPA2基因，LPA2编码一个功能未知蛋白，定位于细胞核与细胞膜。本项目将利用qRT-PCR与组织化学染色法进一步分析LPA2表达模式；通过观察lpa2突变体淀粉颗粒形态及生长素分布差异，分析LPA2在植株重力反应中的作用机制；利用酵母双杂技术、Pull-down、Co-IP及BiFC技术筛选并鉴定LPA2互作蛋白，利用基因超量表达及CRISPR技术研究互作蛋白基因功能；构建双突变体及超表达植株探索LPA2与已知分蘖角度基因的遗传调控关系。通过本项目的实施可以揭示LPA2参与重力反应调控水稻分蘖角度分子机制和调控网络，也可为水稻株型改良提供关键的基因资源，具有重要理论与实践价值。

Rice tiller angle is one of the most important components of rice plant architecture that contributes to determining rice grain yield. The shoot gravitropism largely contributes to the establishment of rice tiller angle. However, the molecular basis of tiller angle has not been well investigated. In our previous study, a loose plant architecture 2 (lpa2) mutant was identified from the EMS mutants of Pingtangheinuo, a black glutinous rice landrance in Guizhou. The lpa2 mutant exhibited loose plant architecture with larger tiller angle and defective shoot gravitropism compared to the wild type. We isolated the LPA2 gene using by map-based cloning approach. LPA2 gene encoded a previously unknown protein that localized in rice nucleus and plasma membrane. To elucidate the critical roles of LPA2 in rice tiller angle, here, we will further examine the expression pattern of LPA2 in different tissues using RT-PCR and GUS histochemical staining.The mode of action of LPA2 by studying the morphology and auxin distribution of lpa2 starch particles will be investigated and analyzed to identify the LPA2-involved mechanism of rice gravitropism. Meanwhile, to reveal the LPA2-involved mechanism in rice tiller angle, we will identify the possible interacting proteins of LPA2 by yeast two-hybrid assay, and combine GST Pull-Down, Co-IP and BiFC assays to confirm the interactions in vitro and in vivo, respectively. Using gene overexpression and CRISPR technology to identify the gene functional of the LPA2 interacting protein. Finally, the genetic relationship between LPA2 and other tiller angle genes will explored by constructing double mutants and overexpressing plants. Based on these results, we will clarify the molecular mechanism and regulatory network of regulating rice shoot gravitropism and tiller angle of LPA2, and also providing resources for genetic improvement of plant architecture.